Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Article
  • Published:

Redirecting cell-type specific cytokine responses with engineered interleukin-4 superkines

Nature Chemical Biology volume 8, pages 990–998 (2012)Cite this article

Subjects

Abstract

Cytokines dimerize their receptors, with the binding of the 'second chain' triggering signaling. In the interleukin (IL)-4 and IL-13 system, different cell types express varying numbers of alternative second receptor chains (γc or IL-13Rα1), forming functionally distinct type I or type II complexes. We manipulated the affinity and specificity of second chain recruitment by human IL-4. A type I receptor–selective IL-4 'superkine' with 3,700-fold higher affinity for γc was three- to ten-fold more potent than wild-type IL-4. Conversely, a variant with high affinity for IL-13Rα1 more potently activated cells expressing the type II receptor and induced differentiation of dendritic cells from monocytes, implicating the type II receptor in this process. Superkines showed signaling advantages on cells with lower second chain numbers. Comparative transcriptional analysis reveals that the superkines induce largely redundant gene expression profiles. Variable second chain numbers can be exploited to redirect cytokines toward distinct cell subsets and elicit new actions, potentially improving the selectivity of cytokine therapy.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Structure-based engineering of IL-4 superkines.
Figure 2: Effect of IL-4 superkines on intracellular signaling.
Figure 3: Modeling of receptor assemblage in response to varying number of second chains.
Figure 4: Functional activities shown by IL-4 and superkines.
Figure 5: Signaling and internalization kinetics of IL-4 and the two superkines in monocytes.
Figure 6: Distinct patterns of cytokine secretion induced by IL-4 and the two superkines in immature and LPS-matured dendritic cells.

Similar content being viewed by others

Accession codes

Primary accessions

Protein Data Bank

References

  1. Leonard, W.J. Type I cytokines and interferons and their receptors. in Fundamental Immunology (ed. Paul, W.) 741–774 (Lippinscott-Raven, Philadelphia, 1999).

  2. Ihle, J.N. Cytokine receptor signalling. Nature 377, 591–594 (1995).

    Article  CAS  Google Scholar 

  3. Stroud, R.M. & Wells, J.A. Mechanistic diversity of cytokine receptor signaling across cell membranes. Sci. STKE 2004, re7 (2004).

    PubMed  Google Scholar 

  4. Wang, X., Lupardus, P., Laporte, S.L. & Garcia, K.C. Structural biology of shared cytokine receptors. Annu. Rev. Immunol. 27, 29–60 (2009).

    Article  Google Scholar 

  5. Cunningham, B.C. et al. Dimerization of the extracellular domain of the human growth hormone receptor by a single hormone molecule. Science 254, 821–825 (1991).

    Article  CAS  Google Scholar 

  6. LaPorte, S.L. et al. Molecular and structural basis of cytokine receptor pleiotropy in the interleukin-4/13 system. Cell 132, 259–272 (2008).

    Article  CAS  Google Scholar 

  7. Junttila, I.S. et al. Tuning sensitivity to IL-4 and IL-13: differential expression of IL-4Rα, IL-13Rα1, and γc regulates relative cytokine sensitivity. J. Exp. Med. 205, 2595–2608 (2008).

    Article  CAS  Google Scholar 

  8. Ito, T. et al. Distinct structural requirements for interleukin-4 (IL-4) and IL-13 binding to the shared IL-13 receptor facilitate cellular tuning of cytokine responsiveness. J. Biol. Chem. 284, 24289–24296 (2009).

    Article  CAS  Google Scholar 

  9. Mueller, T.D., Zhang, J.L., Sebald, W. & Duschl, A. Structure, binding, and antagonists in the IL-4/IL-13 receptor system. Biochim. Biophys. Acta 1592, 237–250 (2002).

    Article  CAS  Google Scholar 

  10. Nelms, K., Keegan, A.D., Zamorano, J., Ryan, J.J. & Paul, W.E. The IL-4 receptor: signaling mechanisms and biologic functions. Annu. Rev. Immunol. 17, 701–738 (1999).

    Article  CAS  Google Scholar 

  11. Kelly-Welch, A.E., Hanson, E.M., Boothby, M.R. & Keegan, A.D. Interleukin-4 and interleukin-13 signaling connections maps. Science 300, 1527–1528 (2003).

    Article  CAS  Google Scholar 

  12. Hage, T., Sebald, W. & Reinemer, P. Crystal structure of the interleukin-4/receptor α chain complex reveals a mosaic binding interface. Cell 97, 271–281 (1999).

    Article  CAS  Google Scholar 

  13. Andrews, A.L., Holloway, J.W., Holgate, S.T. & Davies, D.E. IL-4 receptor α is an important modulator of IL-4 and IL-13 receptor binding: implications for the development of therapeutic targets. J. Immunol. 176, 7456–7461 (2006).

    Article  CAS  Google Scholar 

  14. Andrews, A.L., Holloway, J.W., Puddicombe, S.M., Holgate, S.T. & Davies, D.E. Kinetic analysis of the interleukin-13 receptor complex. J. Biol. Chem. 277, 46073–46078 (2002).

    Article  CAS  Google Scholar 

  15. Heller, N.M. et al. Type I IL-4Rs selectively activate IRS-2 to induce target gene expression in macrophages. Sci. Signal. 1, ra17 (2008).

    Article  Google Scholar 

  16. Kraich, M. et al. A modular interface of IL-4 allows for scalable affinity without affecting specificity for the IL-4 receptor. BMC Biol. 4, 13 (2006).

    Article  Google Scholar 

  17. Wenzel, S., Wilbraham, D., Fuller, R., Getz, E.B. & Longphre, M. Effect of an interleukin-4 variant on late phase asthmatic response to allergen challenge in asthmatic patients: results of two phase 2a studies. Lancet 370, 1422–1431 (2007).

    Article  CAS  Google Scholar 

  18. Boder, E.T. & Wittrup, K.D. Yeast surface display for screening combinatorial polypeptide libraries. Nat. Biotechnol. 15, 553–557 (1997).

    Article  CAS  Google Scholar 

  19. Conrad, D.H. et al. Effect of B cell stimulatory factor-1 (interleukin 4) on Fcɛ and Fcγ receptor expression on murine B lymphocytes and B cell lines. J. Immunol. 139, 2290–2296 (1987).

    CAS  PubMed  Google Scholar 

  20. Siegel, J.P. & Mostowski, H.S. A bioassay for the measurement of human interleukin-4. J. Immunol. Methods 132, 287–295 (1990).

    Article  CAS  Google Scholar 

  21. Gavutis, M., Jaks, E., Lamken, P. & Piehler, J. Determination of the two-dimensional interaction rate constants of a cytokine receptor complex. Biophys. J. 90, 3345–3355 (2006).

    Article  CAS  Google Scholar 

  22. Wong, M.T. et al. Regulation of human TH9 differentiation by type I interferons and IL-21. Immunol. Cell Biol. 88, 624–631 (2010).

    Article  CAS  Google Scholar 

  23. Dauer, M. et al. Mature dendritic cells derived from human monocytes within 48 hours: a novel strategy for dendritic cell differentiation from blood precursors. J. Immunol. 170, 4069–4076 (2003).

    Article  CAS  Google Scholar 

  24. Levin, A.M. et al. Exploiting a natural conformational switch to engineer an interleukin-2 ′superkine′. Nature 484, 529–533 (2012).

    Article  CAS  Google Scholar 

  25. Pillet, A.H. et al. IL-2 induces conformational changes in its preassembled receptor core, which then migrates in lipid raft and binds to the cytoskeleton meshwork. J. Mol. Biol. 403, 671–692 (2010).

    Article  CAS  Google Scholar 

  26. O′Shea, J.J. & Paul, W.E. Mechanisms underlying lineage commitment and plasticity of helper CD4+ T cells. Science 327, 1098–1102 (2010).

    Article  Google Scholar 

  27. Sosman, J.A., Fisher, S.G., Kefer, C., Fisher, R.I. & Ellis, T.M. A phase I trial of continuous infusion interleukin-4 (IL-4) alone and following interleukin-2 (IL-2) in cancer patients. Ann. Oncol. 5, 447–452 (1994).

    Article  CAS  Google Scholar 

  28. Lutz, M.B. et al. Differential functions of IL-4 receptor types I and II for dendritic cell maturation and IL-12 production and their dependency on GM-CSF. J. Immunol. 169, 3574–3580 (2002).

    Article  CAS  Google Scholar 

  29. Creusot, R.J. et al. A short pulse of IL-4 delivered by DCs electroporated with modified mRNA can both prevent and treat autoimmune diabetes in NOD mice. Mol. Ther. 18, 2112–2120 (2010).

    Article  CAS  Google Scholar 

  30. Creusot, R.J. et al. Tissue-targeted therapy of autoimmune diabetes using dendritic cells transduced to express IL-4 in NOD mice. Clin. Immunol. 127, 176–187 (2008).

    Article  CAS  Google Scholar 

  31. Krutzik, P.O. & Nolan, G.P. Fluorescent cell barcoding in flow cytometry allows high-throughput drug screening and signaling profiling. Nat. Methods 3, 361–368 (2006).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

The authors thank J. Gregorio and K. Weiskopf for assistance and the Stanford Human Immune Monitoring Center. This work was supported by US National Institute of Allergy and Infectious Diseases Division of Intramural Research (I.S.J. and W.E.P.), the Finnish Medical Foundation, the Sigrid Juselius Foundation (I.S.J.), the Howard Hughes Medical Institute (K.C.G.), the US National Institutes of Health (NIH) RO1-AI51321 (K.C.G.) and NIH UO1-DK078123 (C.G.F.) and the Stanford Immunology Program Training Grant (D.L.B.).

Author information

Author notes

  1. Ilkka S Junttila, Remi J Creusot, Ignacio Moraga and Darren L Bates: These authors contributed equally to this work.

Authors and Affiliations

  1. Laboratory of Immunology, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, USA

    Ilkka S Junttila & William E Paul

  2. School of Medicine, University of Tampere, Tampere, Finland

    Ilkka S Junttila

  3. Fimlab Laboratories, Tampere, Finland

    Ilkka S Junttila

  4. Department of Medicine, Division of Immunology and Rheumatology, Stanford University School of Medicine, Stanford, California, USA

    Remi J Creusot, Michael T Wong, Paul J Utz & C Garrison Fathman

  5. Howard Hughes Medical Institute, Stanford University School of Medicine, Stanford, California, USA

    Ignacio Moraga, Darren L Bates, Patrick Lupardus & K Christopher Garcia

  6. Department of Molecular and Cellular Physiology, Stanford University School of Medicine, Stanford, California, USA

    Ignacio Moraga, Darren L Bates, Patrick Lupardus & K Christopher Garcia

  7. Department of Structural Biology, Stanford University School of Medicine, Stanford, California, USA

    Ignacio Moraga, Darren L Bates, Patrick Lupardus & K Christopher Garcia

  8. Program in Immunology, Stanford University School of Medicine, Stanford, California, USA

    Ignacio Moraga, Darren L Bates, Patrick Lupardus & K Christopher Garcia

  9. Department of Pathology, Stanford University School of Medicine, Stanford, California, USA

    Michael N Alonso, Megan M Suhoski & Edgar G Engleman

  10. Laboratory of Systems Biology, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, USA

    Martin Meier-Schellersheim

Authors
  1. Ilkka S Junttila
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Remi J Creusot
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Ignacio Moraga
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Darren L Bates
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Michael T Wong
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Michael N Alonso
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Megan M Suhoski
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Patrick Lupardus
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Martin Meier-Schellersheim
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Edgar G Engleman
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. Paul J Utz
    View author publications

    You can also search for this author in PubMed Google Scholar

  12. C Garrison Fathman
    View author publications

    You can also search for this author in PubMed Google Scholar

  13. William E Paul
    View author publications

    You can also search for this author in PubMed Google Scholar

  14. K Christopher Garcia
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

K.C.G. conceived the project, designed approaches for engineering of IL-4 and initiated subsequent cellular and functional experiments. D.L.B. and P.L. performed protein engineering and biophysical experiments. I.S.J., R.J.C., I.M. and W.E.P. designed and performed signaling experiments. R.J.C., M.M.-S. and I.M. performed transcriptional analysis and Luminex experiments. W.E.P., I.S.J. and M.M.S. performed mathematical modeling using Matlab. M.T.W., M.N.A., M.M.S. and I.M. performed dendritic cell experiments. I.S.J., R.J.C., I.M., D.L.B., C.G.F., P.J.U., W.E.P. and K.C.G. analyzed the data. P.J.U. and E.G.E. provided reagents and guidance for human primary cell experiments. I.S.J., R.J.C., I.M., W.E.P. and K.C.G. wrote the manuscript.

Corresponding author

Correspondence to K Christopher Garcia.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Text and Figures

Supplementary Methods and Supplementary Results (PDF 7551 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Junttila, I., Creusot, R., Moraga, I. et al. Redirecting cell-type specific cytokine responses with engineered interleukin-4 superkines. Nat Chem Biol 8, 990–998 (2012). https://doi.org/10.1038/nchembio.1096

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nchembio.1096

This article is cited by

Search

Advanced search

Quick links

Nature Briefing: Translational Research

Sign up for the Nature Briefing: Translational Research newsletter — top stories in biotechnology, drug discovery and pharma.

Get what matters in translational research, free to your inbox weekly. Sign up for Nature Briefing: Translational Research